Detection of circulating hepatocellular carcinoma cells in peripheral venous blood by reverse transcription-polymerase chain reaction

Objective: To detect circulating hepatocellular carcino-ma by demonstrating hepatocellular carcinoma cells orhepatocyte-associated mRNA in the nuclear cell com-ponent of peripheral blood (PBL).Methods: Peripheral blood (5 ml) samples were ob-tained from 93 patients with hepatocellular carcinoma(HCC) and from 33 control subjects (9 with liver cir-rhosis after hepatitis B,14 with chronic hepatitis B,10with normal liver function). To identify HCC cells inperipheral blood, liver-specific human alpha-fetopro-tein (AFP) mRNA was amplified from total RNA ex-tracted from whole blood by reverse transcription-polymerase chain reaction.Results: AFPmRNA was detected in 50 blood samplesfrom the HCC patients (50/93, 53.8%). In contrast,there were no clinical control patients whose samplesshowed detectable AFPmRNA in PBL. The presence ofAFPmRNA in blood seemed to be correlated with thestage (by TNM classification) of HCC, the serum AFPvalue, and the presence of intrahepatic metastasis,portal vein thrombosis, tumor diameter and/or distantmetastasis. In addition, AFPmRNA was detected in theblood of 21 patients with metastasis at extrahepaticorgans (100%) in contrast to 29 (40.3%)of 72 pa-tients without metastasis.Conclusion: The presence of AFPmRNA in peripheralblood may be an indicator of malignant hepatocytes,which might predict hematogenous spreading metasta-sis of tumor cells in patients with HCC.

Apoptosis in acute rejection of hamster-to-rat liver transplantation

Objective: To investigate whether apoptotic cell death is involved in liver xenograft rejection and the molec- ular mechanism of apoptosis. Methods: After hamster-to-rat orthotopic liver trans- plantation, apoptosis in the xenograft was observed histologically and by in situ end-labelling of fragmen- ted DNA. CD_8 antigen, perforin, Fas-L and TGF-β_1 were observed immunohistochemically in the graft. Results: In xenogenic rejection, OX_8 (CD_8) positive T lymphocyte was observed on day 2 post-transplan- tation, evidently on day 5. The expression of perfor- in and Fas-L in grafts occurred on day 4 post-trans- plantation, and it was more evident in the rejection. In control group, the lymphocyte was rarely seen, and the expression of Fas-L and perforin was not found. Both xenogeneic and syngeneic grafts showed the expression of TGF-β_1 on the first day after trans- plantation. The expression of TGF-β_1, however, in- creased subsequently in the xenogeneic graft in con- trast to its normalization in the syngeneic graft. In the xenogeneic graft, apoptosis occurred on day 1 af- ter transplantation, decreased on day 2, increased on day 3, and peaked on day 5. Apoptosis was similar in the syngeneic and xenogeneic grafts on day 1 after transplantation, but decreased to normal one day lat- er. The more severe apoptosis, the more severe acute rejection, and the more evident expression of perfor- in, Fas-L and TGF-β_1. Conclusion: Apoptosis as a mechanism of cell death exists in the acute rejection of liver xenograft, and it is related closely to the expression of perforin, TGF- β_1 and Fas-L.

Nested RT-PCR in detection of blood AFPmRNA in animal model of human hepatocellular carcinoma

OBJECTIVES: To detect blood AFPmRNA in the nude mice bearing with human hepatocellular carcinoma (HCC) by using nested reverse transcriptase polymerase chain reaction (nested RT-PCR), and assess its significance in HCC distant metastasis. METHOD: We detected 20 blood samples from the nude mice bearing with human HCC by nested RT-PCR to find out AFPmRNA. RESULT: AFPmRNA was detected in 6 blood samples from the nude mice hearing with human HCC (30. 0%), in which 4 mice developed distant metastasis. CONCLUSION: AFPmRNA may be used as an efficient and sensitive marker to detect blood spread of HCC cells. It can predict the occurrence of HCC distant metastasis.

Changes of AFPmRNA in blood after percutaneous ethanol injection treatment of hepatocellular carcinoma

Objective: To study the effects of percutaneous etha- nol injection (PEI) in treatment of hepatocellular carcinoma (HCC). Methods: Blood samples were taken from 9 HCC pa- tients and detected by nested RT-PCR to find out AFPmRNA after PEI treatment 3-5 times within 2 weeks. Results: Three (44.44%) of 4 patients with AFPmR- NA positive turned to be negative after PEI treatment and 1 with AFPmRNA positive (11.11%) lasted within 2 weeks. Conclusion: PEI may diminish the possibility of dis- tant metastasis of HCC.

Expression of AFPmRAN in blood of patients with recurrent hepatocellular carcinoma

OBJECTIVES: To detect the expression of human alpha-fetoprotein mRNA (AFPmRNA) in peripheral blood (PBL) of recurrent hepatocellular carcinoma (HCC) by using nested reverse transcriptase polymerase chain reaction (nested RT-PCR) and to discuss relations between AFPmRNA and recurrent HCC. METHODS: Five ml peripheral blood samples were obtained from 19 patients with recurrent HCC. To identify HCC cells in PBL, liver-specific AFPmRNA was amplified from the total RNA extracted from the whole blood by nested RT-PCR. RESULTS: Seven patients (36.8%) with recurrent HCC were detected AFPmRNA, and 12 patients with recurrent HCC were AFPmRNA negative (63.2%). There was no significant relation between patients with AFPmRNA positive and AFPmRNA negative (P>0.05) in judging the origin of recurrent HCC. CONCLUSION: AFPmRNA may not be used as a marker of origin of recurrent HCC.